MECHANISMS OF MULTIDRUG RESISTANCE IN PSEUDOMONAS AERUGINOSA CLINICAL ISOLATES FROM COMPANION ANIMAL AND HUMAN PATIENTS

Abstract

Pseudomonas aeruginosa has been notoriously known for its intrinsic and acquired resistance to multiple antimicrobial agents simultaneously. So far, mechanisms underlining multidrug resistance (MDR) in P. aeruginosa have been extensively studied but mostly, in laboratory stains. Such information is still limited in the clinical isolates. Therefore, the goal of this dissertation was to understand genetic mechanisms responsible for MDR phenotype in P. aeruginosa clinical isolates. We examined two major MDR mechanisms, including integrons and multidrug efflux systems (Mex) of the Resistance-Nodulation-Cell Division (RND) family, in the P. aeruginosa clinical isolates from animals and humans (n=114). One of the major findings was the high prevalence of class 1 integrons containing resistance genes cassette (69.3%) in the P. aeruginosa human isolates admitted to a hospital (n=101). Five novel resistance gene cassettes including aacA7-cmlA, aadB-blaOXA-10-aadA1, aadB-arr-2-cmlA- blaOXA-10-aadA1, aadB-cmlA-aadA1 and aadB-cmlA- blaOXA-10-aadA15 were identified (50.5%). Up to date, there are up to 12 RND efflux operons found on P. aeruginosa chromosome, of which six Mex systems (i.e. MexAB-OprM, MexCD-OprJ, MexEF-OprN, MexJK, MexVW and MexXY) have been shown to extrude antimicrobial substrates and play a role in cross-resistance among antimicrobials. The predominant finding was the simultaneous expression of at least 3 Mex systems in a single P. aeruginosa isolate from either humans or animals (n=43), suggesting that expression of several Mex systems concomitantly contributes to multiple drug resistance in the clinical isolates. Role of normally silent Mex systems including MexCD-OprJ, MexEF-OprN, MexJK and MexVW were highlighted. Method for simultaneous detection of four-clinically important Mex systems were developed using combination of resistance-phenotypic markers and multiplex RT-PCR. In conclusion, the results demonstrated an intriguing and complex picture of expression and regulation of the Mex systems in the P. aeruginosa clinical isolates. The observations demonstrated that multiple pathways existing to participate in MDR phenotypes and contribution of normally silent Mex systems should not be underestimated. In addition, control/prevention strategies for antimicrobial resistance need to be encouraged e.g. responsible therapeutic use of antimicrobials, routine antimicrobial resistance monitoring and public education.